Humans exhibit an innate immunity against the mammalian parasite Trypanosoma brucei brucei, the causative agent of Nagana in African cattle. The morphologically indistinct T. brucei subspecies T. b. rhodesiense and T. b. gambiense are responsible for fatal sleeping sickness in humans, a disease of epidemic proportions in sub-Saharan Africa. Immunity against T. b. brucei is mediated by a subset of high-density lipoprotein termed trypanosome lytic factor. Susceptible trypanosomes are killed in human plasma when trypanosome lytic factor is endocytosed and trafficked to the lysosome. The acidic environment of the lysosome facilitates a membrane disrupting activity by the high-density lipoprotein particle that ultimately results in cell death. The goal of this project is to define the mechanism of membrane disruption by trypanosome lytic factor (Specific Aim I) and elucidate the role of an unusual retained hydrophobic N-terminal signal peptide in haptoglobin related protein (a component of trypanosome lytic factor) (Specific Aim II). To this end we have developed a model membrane system to test the hypothesis that trypanosome lytic factor disrupts membranes through stepwise binding, stable association and protein mediated lipid peroxidation and pore formation. We will also test the hypothesis that the retained signal peptide is necessary and sufficient for assembly of haptoglobin related protein and that it mediates a trypanolytic activity by this protein. PUBLIC HEALTH RELEVANCE: A thorough understanding of the mechanism by which TLF targets and disrupts lysosomal membranes may inform the development of therapeutic agents against not only pathogenic trypanosomes but other pathogenic cells in general. Additionally, examining the role of the signal peptide will contribute to a fundamental understanding of the biology of HDL.